Alterazione di risposte fisiologiche nei mitili Mytilus galloprovincialis esposti a caffeina, contaminante emergente dell'ambiente marino.

L’elevata presenza dei residui dei farmaci nell’ambiente acquatico desta preoccupazione per la salute della fauna acquatica e per un eventuale rischio per l’uomo. Dopo l’assunzione, i farmaci vengono escreti come tali o come metaboliti attivi, risultando spesso resistenti ai processi di trattamento delle acque. Per questo motivo alcuni di essi sono pseudo-persistenti raggiungendo concentrazioni di ng-µg/L. I farmaci sono molecole disegnate per essere biologicamente attive a basse concentrazioni su bersagli specifici, per questo motivo possono indurre effetti anche su specie non target con bersagli molecolari simili all’uomo a concentrazioni ambientali. A tal proposito il presente lavoro intende investigare gli effetti della caffeina, ampiamente usata come costituente di bevande e come farmaco, presente nelle acque superficiali a concentrazioni di ng-µg/L. Come organismo di studio è stato scelto il Mytilus galloprovincialis, sfruttando le conoscenze disponibili in letteratura circa le sue risposte ai contaminanti ambientali. I mitili sono stati esposti in acquario a caffeina (5, 50 e 500 ng/L) per 7 giorni e poi analizzati attraverso una batteria di otto biomarker, alterazioni fisiologiche o biochimiche che forniscono informazioni circa lo stato di salute degli animali. I metodi utilizzati sono stati diversi a seconda dei biomarker (analisi citochimiche e saggi enzimatici). Le concentrazioni sono state scelte nel range ambientale. L’esposizione ha prodotto alterazioni della stabilità della membrana lisosomiale negli emociti e l’instaurarsi di processi di detossificazione nella ghiandola digestiva, evidenziati dall’aumento dell’attività della glutatione S-transferasi. Gli altri biomarker non mettono in evidenza che la caffeina, in queste condizioni sperimentali, possa indurre alterazioni della funzionalità lisosomiale, effetti ossidativi o neurotossici. I dati ottenuti sui mitili, quindi, suggeriscono che la caffeina, anche nel range di concentrazioni ambientali più elevato, possa essere considerata un contaminante che desta bassa preoccupazione.

Brain-Computer Interfaces for augmented communication: Asynchronous and adaptive algorithms and evaluation with end users

This thesis aimed at addressing some of the issues that, at the state of the art, avoid the P300-based brain computer interface (BCI) systems to move from research laboratories to end users’ home. An innovative asynchronous classifier has been defined and validated. It relies on the introduction of a set of thresholds in the classifier, and such thresholds have been assessed considering the distributions of score values relating to target, non-target stimuli and epochs of voluntary no-control. With the asynchronous classifier, a P300-based BCI system can adapt its speed to the current state of the user and can automatically suspend the control when the user diverts his attention from the stimulation interface. Since EEG signals are non-stationary and show inherent variability, in order to make long-term use of BCI possible, it is important to track changes in ongoing EEG activity and to adapt BCI model parameters accordingly. To this aim, the asynchronous classifier has been subsequently improved by introducing a self-calibration algorithm for the continuous and unsupervised recalibration of the subjective control parameters. Finally an index for the online monitoring of the EEG quality has been defined and validated in order to detect potential problems and system failures. This thesis ends with the description of a translational work involving end users (people with amyotrophic lateral sclerosis-ALS). Focusing on the concepts of the user centered design approach, the phases relating to the design, the development and the validation of an innovative assistive device have been described. The proposed assistive technology (AT) has been specifically designed to meet the needs of people with ALS during the different phases of the disease (i.e. the degree of motor abilities impairment). Indeed, the AT can be accessed with several input devices either conventional (mouse, touchscreen) or alterative (switches, headtracker) up to a P300-based BCI.

Effects of temperature and acidity on Mediterranean benthic calcifiers, in the face of globale climate change

The Mediterranean Sea is expected to react faster to global change compared to the ocean and is already showing more pronounced warming and acidification rates. A study performed along the Italian western coast showed that porosity of the skeleton increases with temperature in the zooxanthellate (i.e. symbiotic with unicellular algae named zooxanthellae) solitary scleractinian Balanophyllia europaea while it does not vary with temperature in the solitary non-zooxanthellate Leptopsammia pruvoti. These results were confirmed by another study that indicated that the increase in porosity was accompanied by an increase of the fraction of the largest pores in the pore-space, perhaps due to an inhibition of the photosynthetic process at elevated temperatures, causing an attenuation of calcification. B. europaea, L. pruvoti and the colonial non-zooxanthellate Astroides calycularis, transplanted along a natural pH gradient, showed that high temperature exacerbated the negative effect of lowered pH on their mortality rates. The growth of the zooxanthellate species did not react to reduced pH, while the growth of the two non-zooxanthellate species was negatively affected. Reduced abundance of naturally occurring B. europaea, a mollusk, a calcifying and a non-calcifying macroalgae were observed along the gradient while no variation was seen in the abundance of a calcifying green alga. With decreasing pH, the mineralogy of the coral and mollusk did not change, while the two calcifying algae decreased the content of aragonite in favor of the less soluble calcium sulphates and whewellite (calcium oxalate), possibly as a mechanism of phenotypic plasticity. Increased values of porosity and macroporosity with CO2 were observed in B. europaea specimens, indicating reduces the resistance of its skeletons to mechanical stresses with increasing acidity. These findings, added to the negative effect of temperature on various biological parameters, generate concern on the sensitivity of this zooxanthellate species to the envisaged global climate change scenarios.

Identification of stock units of Mullus barbatus (Linnaeus, 1758) in the Italian seas: development of genomic 2b-RAD markers and biological variation analysis.

This thesis is settled within the STOCKMAPPING project, which represents one of the studies that were developed in the framework of RITMARE Flagship project. The main goals of STOCKMAPPING were the creation of a genomic mapping for stocks of demersal target species and the assembling of a database of population genomic, in order to identify stocks and stocks boundaries. The thesis focuses on three main objectives representing the core for the initial assessment of the methodologies and structure that would be applied to the entire STOCKMAPPING project: individuation of an analytical design to identify and locate stocks and stocks boundaries of Mullus barbatus, application of a multidisciplinary approach to validate biological methods and an initial assessment and improvement for the genotyping by sequencing technique utilized (2b-RAD). The first step is the individuation of an analytical design that has to take in to account the biological characteristics of red mullet and being representative for STOCKMAPPING commitments. In this framework a reduction and selection steps was needed due to budget reduction. Sampling areas were ranked according the individuation of four priorities. To guarantee a multidisciplinary approach the biological data associated to the collected samples were used to investigate differences between sampling areas and GSAs. Genomic techniques were applied to red mullet for the first time so an initial assessment of molecular protocols for DNA extraction and 2b-RAD processing were needed. At the end 192 good quality DNAs have been extracted and eight samples have been processed with 2b-RAD. Utilizing the software Stacks for sequences analyses a great number of SNPs markers among the eight samples have been identified. Several tests have been performed changing the main parameter of the Stacks pipeline in order to identify the most explicative and functional sets of parameters.

Valutazione di marker diagnostici, epidemiologia e nuove prospettive terapeutiche nell' avvelenamento da rodenticidi anticoagulanti nel cane.

In Medicina Veterinaria l'avvelenamento da rodenticidi anticoagulanti è conosciuto e studiato ormai da anni, essendo una delle intossicazioni più comunemente riscontrate nelle specie non target. In letteratura si rinvengono numerose pubblicazioni ma alcuni aspetti sono rimasti ancora inesplorati.Questo studio si propone di valutare il processo infiammatorio, mediante le proteine di fase acuta (APPs), in corso di fenomeni emorragici, prendendo come modello reale un gruppo di soggetti accidentalmente avvelenati da rodenticidi anticoagulanti. I 102 soggetti avvelenati presentano un valore più elevato di proteina C reattiva (CRP)con una mediana di 4.77 mg/dl statisticamente significativo rispetto alla mediana delle due popolazioni di controllo di pari entità numerica create con cross match di sesso, razza ed età; rispettivamente 0.02 mg/dl dei soggetti sani e 0.37 mg/dl dei soggetti malati di altre patologie. Inoltre all'interno del gruppo dei soggetti avvelenati un valore di CRP elevato all'ammissione può predisporre al decesso. La proteina C reattiva assume quindi un ruolo diagnostico e prognostico in questo avvelenamento. Un'altra finalità, di non inferiore importanza, è quella di definire una linea guida terapeutica con l'ausilio di biomarker coagulativi e di valutare la sicurezza della vitamina K per via endovenosa: in 73 cani, non in terapia con vitamina k, intossicati da rodenticidi anticoagulanti, i tempi della coagulazione (PT ed aPTT) ritornano nel range di normalità dopo 4 ore dalla prima somministrazione di 5 mg/kg di vitamina k per via endovenosa e nessun soggetto durante e dopo il trattamento ha manifestato reazioni anafilattiche, nessuno dei pazienti ha necessitato trasfusione ematica e tutti sono sopravvissuti. Infine si è valutata l'epidemiologia dell'ingestione dei prodotti rodenticidi nella specie oggetto di studio e la determinazione dei principi attivi mediante cromatografia liquida abbinata a spettrofotometria di massa (UPLC-MS/MS).

Development and application of mass-spectrometric methods for the quantification and characterization of organic compounds in ice cores

Eisbohrkerne stellen wertvolle Klimaarchive dar, da sie atmosphärisches Aerosol konservieren. Die Analyse chemischer Verbindungen als Bestandteil atmosphärischer Aerosole in Eisbohrkernen liefert wichtige Informationen über Umweltbedingungen und Klima der Vergangenheit. Zur Untersuchung der α-Dicarbonyle Glyoxal und Methylglyoxal in Eis- und Schneeproben wurde eine neue, sensitive Methode entwickelt, die die Stir Bar Sorptive Extraction (SBSE) mit der Hochleistungsflüssigchromatographie-Massenspektrometrie (HPLC-MS) kombiniert. Zur Analyse von Dicarbonsäuren in Eisbohrkernen wurde eine weitere Methode entwickelt, bei der die Festphasenextraktion mit starkem Anionenaustauscher zum Einsatz kommt. Die Methode erlaubt die Quantifizierung aliphatischer Dicarbonsäuren (≥ C6), einschließlich Pinsäure, sowie aromatischer Carbonsäuren (wie Phthalsäure und Vanillinsäure), wodurch die Bestimmung wichtiger Markerverbindungen für biogene und anthropogene Quellen ermöglicht wurde. Mit Hilfe der entwickelten Methoden wurde ein Eisbohrkern aus den Schweizer Alpen analysiert. Die ermittelten Konzentrationsverläufe der Analyten umfassen die Zeitspanne von 1942 bis 1993. Mittels einer Korrelations- und Hauptkomponentenanalyse konnte gezeigt werden, dass die organischen Verbindungen im Eis hauptsächlich durch Waldbrände und durch vom Menschen verursachte Schadstoffemissionen beeinflusst werden. Im Gegensatz dazu sind die Konzentrationsverläufe einiger Analyten auf den Mineralstaubtransport auf den Gletscher zurückzuführen. Zusätzlich wurde ein Screening der Eisbohrkernproben mittels ultrahochauflösender Massenspektrometrie durchgeführt. Zum ersten Mal wurden in diesem Rahmen auch Organosulfate und Nitrooxyorganosulfate in einem Eisbohrkern identifiziert.

Methodenentwicklung zur Spurenanalyse organischer Biomarker in Speläothemen und Tropfwasser mittels Flüssigchromatographie-(HR-)Massenspektrometrie

Speläotheme und Tropfwasser spielen eine wichtige Rolle bei der Erforschung von Naturerscheinungen. Während sich bisherige Studien größtenteils auf anorganische Proxies konzentrieren, wächst das Interesse an organischen Biomarkern, vor allem Lipidbiomarkern.rnAufgrund dessen wurde eine neue Methode entwickelt, um Fettsäuren mit einer geradzahligen Kettenlänge C12-C20 in Speläothem- und Tropfwasserproben zu bestimmen. Dabei kam eine Festphasenextraktion mit anschließender HPLC-MS Messung zum Einsatz. Die Methode wurde auf mehrere Proben aus der Herbstlabyrinth-Adventhöhle angewendet. Die benötigte Probenmenge wurde im Vergleich zu früheren Studien von etwa 4 L auf 60-100 mL bei Tropfwasser und von 10-100 g auf ca. 0,5-3,5 g bei Stalagmitproben reduziert. Es konnte eine interne Variation der Analyten festgestellt werden. Korrelationen der Fettsäuren ließen vermuten, dass C12-C18 von der gleichen Quelle stammen, während C20 teilweise andere Quellen besitzt. Korrelationen mit δ13C verdeutlichten, dass es einen Zusammenhang zwischen der Vegetationsdichte und dem Vorkommen der Fett-säuren in dem Probenmaterial gibt. Vergleiche mit Mg Konzentrationen zeigten, dass die Niederschlagsmenge zwar den Transport der Fettsäuren mit dem Tropfwasser beeinflusst, allerdings nicht ihre ermittelte Konzentration in Stalagmitproben. Die ermittelten Ergebnisse ließen darauf schließen, dass eindeutigere Vegetations-Proxies als die Fettsäuren gefunden werden müssen. Ein erstmaliges non-target screening verdeutlichte, dass Lignine und Tannine als charakteristische Biomarker eingesetzt werden können.rn

Temporal variation of genetic diversity and differentiation of albacore, Thunnus alalunga, in the Mediterranean Sea

This study is on albacore (Thunnus alalunga, Bonnaterre 1788), an epi- and mesopelagic oceanic tuna species cosmopolitan in the tropical and temperate waters of all oceans including the Mediterranean Sea, extending in a broad band between 40°N and 40°S. What it’s known about albacore population structure is based on different studies that used fisheries data, RFLP, mtDNA control region and nuDNA markers, blood lectins analysis, individual tags and microsatellite. At the moment, for T. alalunga six management units are recognized: the North Pacific, South Pacific, Indian, North Atlantic, South Atlantic and Mediterranean stocks. In this study I have done a temporal and spatial comparison of genetic variability between different Mediterranean populations of Thunnus alalunga matching an historical dataset ca. from 1920s composed of 43 individuals divided in 3 populations (NADR, SPAIN and CMED) with a modern dataset composed of 254 individuals and 7 populations (BAL, CYP, LIG, TYR, TUR, ADR, ALB). The investigation was possible using a panel of 94 nuclear SNPs, built specifically for the target species at the University of Basque Country UPV/EHU. First analysis done was the Hardy-Weinberg, then the number of clusters (K) was determined using STRUCTURE and to assess the genetic variability, allele frequencies, the average number of alleles per locus, expected (He) and observed (Ho) heterozygosis, and the index of polymorphism (P) was used the software Genetix. Historical and modern samples gives different results, showing a clear loss of genetic diversity over time leading to a single cluster in modern albacore instead of the two found in historical samples. What this study reveals is very important for conservation concerns, and additional research endeavours are needed.

Are there genetic breaks between Atlantic and Pacific Yellowfin tuna (Thunnus albacares) populations? A preliminary study based on microsatellites gene variation

Yellowfin tuna (Thunnus albacares, YFT, Bonnaterre 1788) is one of the most important market tuna species in the world. The high mortality of juveniles is in part caused by their bycatch. Indeed, if unregulated, it could permanently destabilize stocks health. For this reason investigating and better knowing the stock boundaries represent a crucial concern. Aim of this thesis was to preliminary investigate the YFT population structure within and between Atlantic and Pacific Oceans through the analysis of genetic variation at eight microsatellite loci and assess the occurrence of barriers to the gene flow between Oceans. For this propouse we collected 4 geographical samples coming from Atlantic and Pacific Ocean and selected a panel of 8 microsatellites loci developped by Antoni et al., (2014). Samples 71-2-Y and 77-2-Y, came from rispectively west central pacific ocean (WCPO) and east central pacific ocean (ECPO), instead samples 41-1-Y and 34-2-Y derive from west central atlantic ocean (WCAO) and east central atlantic ocean (ECAO). Total 160 specimens were analyzed (40 per sample) and were carried out several genetic information as allele frequencies, allele number, allelic richness, HWE (using He and Ho) and pairwise Fst genetic distance. Results obtained, may support the panmictic theory of this species, only one of pairwise Fst obtained is statistically significant (Fst= 0.00927; pV= 0.00218) between 41-1-Y and 71-2-Y samples. Results suggest low genetic differentiation and consequent high level of gene flow between Atlantic and Pacific populations. Furthermore, we performed an analysis of molecular taxonomy through the use of ATCO (the flaking region between ATPse6 and cytochrome oxidase subunit III genes mt DNA, to discriminate within the gener Thunnus two of the related species (Yellofin and bigeye tuna) according with their difficult recognition at certain size (<40 cm). ATCO analysis in this thesis, has provided strong discriminate evidence between the target species proving to be one of the most reliable genetic tools capable to indagate within the genus Thunnus. Thus, our study has provided useful information for possible use of this protocol for conservation plans and management of this fish stocks.

Citizen science as a tool for the environmental quality assessment of the Mediterranean coastal habitats

The growing need to assess the environmental status of the Mediterranean coastal marine habitats and the large availability of data collected by Reef Check Italia onlus (RCI) volunteers suggest the possibility to develop innovative and reliable indices that may support decision makers in applying conservation strategies. The aims of this study were to check the reliability of data collected by RCI volunteers, analyse the spatial and temporal distribution of RCI available data, resume the knowledge on the biology and ecology of the monitored species, and develop innovative indices to asses the ecological quality of Mediterranean subtidal rocky shores and coralligenous habitats. Subtidal rocky shores and coralligenous were chosen because these are the habitats more attractive for divers; therefore mlst data are referring to them, moreover subtidal rocky bottom are strongly affected by coastal urbanisation, land use, fishing and tourist activities, that increase pollution, turbidity and sedimentation. Non-indigenous species (NIS) have been recognized as a major threat to the integrity of Mediterranean native communities because of their proliferation, spread and impact on resident communities. Monitoring of NIS’ spreading dynamics at the basin spatial scale is difficult but urgent. According to a field test, the training provided by RCI appears adequate to obtain reliable data by volunteers. Based on data collected by RCI volunteers, three main categories of indices were developed: indices based on species diversity, indices on the occurrence non-indigenous species, and indices on species sensitive toward physical, chemical and biological disturbances. As case studies, indices were applied to stretches of coastline defined according to management criteria (province territories and marine protected areas). The assessments of ecological quality in the Tavolara Marine Protected Area using the species sensitivities index were consisten with those previously obtained with traditional methods.

No adverse effect of genetically modified antifungal wheat on decomposition dynamics and the soil fauna community - a field study

The cultivation of genetically modified (GM) plants has raised several environmental concerns. One of these concerns regards non-target soil fauna organisms, which play an important role in the decomposition of organic matter and hence are largely exposed to GM plant residues. Soil fauna may be directly affected by transgene products or indirectly by pleiotropic effects such as a modified plant metabolism. Thus, ecosystem services and functioning might be affected negatively. In a litterbag experiment in the field we analysed the decomposition process and the soil fauna community involved. Therefore, we used four experimental GM wheat varieties, two with a race-specific antifungal resistance against powdery mildew (Pm3b) and two with an unspecific antifungal resistance based on the expression of chitinase and glucanase. We compared them with two non-GM isolines and six conventional cereal varieties. To elucidate the mechanisms that cause differences in plant decomposition, structural plant components (i.e. C:N ratio, lignin, cellulose, hemicellulose) were examined and soil properties, temperature and precipitation were monitored. The most frequent taxa extracted from decaying plant material were mites (Cryptostigmata, Gamasina and Uropodina), springtails (Isotomidae), annelids (Enchytraeidae) and Diptera (Cecidomyiidae larvae). Despite a single significant transgenic/month interaction for Cecidomyiidae larvae, which is probably random, we detected no impact of the GM wheat on the soil fauna community. However, soil fauna differences among conventional cereal varieties were more pronounced than between GM and non-GM wheat. While leaf residue decomposition in GM and non-GM wheat was similar, differences among conventional cereals were evident. Furthermore, sampling date and location were found to greatly influence soil fauna community and decomposition processes. The results give no indication of ecologically relevant adverse effects of antifungal GM wheat on the composition and the activity of the soil fauna community.

Monitoring Butterfly Abundance: Beyond Pollard Walks

Most butterfly monitoring protocols rely on counts along transects (Pollard walks) to generate species abundance indices and track population trends. It is still too often ignored that a population count results from two processes: the biological process (true abundance) and the statistical process (our ability to properly quantify abundance). Because individual detectability tends to vary in space (e.g., among sites) and time (e.g., among years), it remains unclear whether index counts truly reflect population sizes and trends. This study compares capture-mark-recapture (absolute abundance) and count-index (relative abundance) monitoring methods in three species (Maculinea nausithous and Iolana iolas: Lycaenidae; Minois dryas: Satyridae) in contrasted habitat types. We demonstrate that intraspecific variability in individual detectability under standard monitoring conditions is probably the rule rather than the exception, which questions the reliability of count-based indices to estimate and compare specific population abundance. Our results suggest that the accuracy of count-based methods depends heavily on the ecology and behavior of the target species, as well as on the type of habitat in which surveys take place. Monitoring programs designed to assess the abundance and trends in butterfly populations should incorporate a measure of detectability. We discuss the relative advantages and inconveniences of current monitoring methods and analytical approaches with respect to the characteristics of the species under scrutiny and resources availability.

[(99m)Tc]Demotate 2 in the detection of sst(2)-positive tumours: a preclinical comparison with [(111)In]DOTA-tate

PURPOSE: The aim of this study was to evaluate [(99m)Tc]Demotate 2 ([(99m)Tc-N(4) (0-1),Asp(0),Tyr(3)]octreotate) as a candidate for in vivo imaging of sst(2)-positive tumours and to compare it with [(111)In]DOTA-tate ([(111)In-DOTA(0),Tyr(3)]octreotate). METHODS: Labelling of Demotate 2 with (99m)Tc was performed at room temperature using SnCl(2) as reductant in the presence of citrate at alkaline pH. Radiochemical analysis involved ITLC and HPLC methods. Peptide conjugate affinities for sst(2) were determined by receptor autoradiography on rat brain cortex sections using [DOTA(0),(125)I-Tyr(3)]octreotate as the radioligand. The affinity profile of Demotate 2 for human sst(1)-sst(5) was studied by receptor autoradiography in cell preparations using the universal somatostatin radioligand [(125)I][Leu(8),(D: )Trp(22),Tyr(25)]somatostatin-28. The internalisation rates of [(99m)Tc]Demotate 2 and [(111)In]DOTA-tate were compared in sst(2)-positive and -negative control cell lines. Biodistribution of radiopeptides was studied in male Lewis rats bearing CA20948 tumours. RESULTS: Peptide conjugates showed selectivity and a high affinity binding for sst(2) (Demotate 2 IC(50)=3.2 nM and DOTA-tate IC(50)=5.4 nM). [(99m)Tc]Demotate 2, like [(111)In]DOTA-tate, internalised rapidly in all sst(2)-positive cells tested, but not in sst(2)-negative control cells. After injection in CA20948 tumour-bearing rats both radiopeptides showed high and specific uptake in the sst(2)-positive organs and in the implanted tumour and rapid excretion from non-target tissues via the kidneys. CONCLUSION: [(99m)Tc]Demotate 2, similarly to the known sst(2)-targeting agent [(111)In]DOTA-tate, showed promising biological qualities for application in the scintigraphy of sst(2)-positive tumours.

A randomized, blinded, multicenter trial of lipid-associated amphotericin B alone versus in combination with an antibody-based inhibitor of heat shock protein 90 in patients with invasive candidiasis

BACKGROUND: Mycograb (NeuTec Pharma) is a human recombinant monoclonal antibody against heat shock protein 90 that, in laboratory studies, was revealed to have synergy with amphotericin B against a broad spectrum of Candida species. METHODS: A double-blind, randomized study was conducted to determine whether lipid-associated amphotericin B plus Mycograb was superior to amphotericin B plus placebo in patients with culture-confirmed invasive candidiasis. Patients received a lipid-associated formulation of amphotericin B plus a 5-day course of Mycograb or placebo, having been stratified on the basis of Candida species (Candida albicans vs. non-albicans species of Candida). Inclusion criteria included clinical evidence of active infection at trial entry plus growth of Candida species on culture of a specimen from a clinically significant site within 3 days after initiation of study treatment. The primary efficacy variable was overall response to treatment (clinical and mycological resolution) by day 10. RESULTS: Of the 139 patients enrolled from Europe and the United States, 117 were included in the modified intention-to-treat population. A complete overall response by day 10 was obtained for 29 (48%) of 61 patients in the amphotericin B group, compared with 47 (84%) of 56 patients in the Mycograb combination therapy group (odds ratio [OR], 5.8; 95% confidence interval [CI], 2.41-13.79; P<.001). The following efficacy criteria were also met: clinical response (52% vs. 86%; OR, 5.4; 95% CI, 2.21-13.39; P<.001), mycological response (54% vs. 89%; OR, 7.1; 95% CI, 2.64-18.94; P<.001), Candida-attributable mortality (18% vs. 4%; OR, 0.2; 95% CI, 0.04-0.80; P = .025), and rate of culture-confirmed clearance of the infection (hazard ratio, 2.3; 95% CI, 1.4-3.8; P = .001). Mycograb was well tolerated. CONCLUSIONS: Mycograb plus lipid-associated amphotericin B produced significant clinical and culture-confirmed improvement in outcome for patients with invasive candidiasis.

DOTA-PESIN, a DOTA-conjugated bombesin derivative designed for the imaging and targeted radionuclide treatment of bombesin receptor-positive tumours

PURPOSE: We aimed at designing and developing a novel bombesin analogue, DOTA-PEG(4)-BN(7-14) (DOTA-PESIN), with the goal of labelling it with (67/68)Ga and (177)Lu for diagnosis and radionuclide therapy of prostate and other human cancers overexpressing bombesin receptors. METHODS: The 8-amino acid peptide bombesin (7-14) was coupled to the macrocyclic chelator DOTA via the spacer 15-amino-4,7,10,13-tetraoxapentadecanoic acid (PEG(4)). The conjugate was complexed with Ga(III) and Lu(III) salts. The GRP receptor affinity and the bombesin receptor subtype profile were determined in human tumour specimens expressing the three bombesin receptor subtypes. Internalisation and efflux studies were performed with the human GRP receptor cell line PC-3. Xenografted nude mice were used for biodistribution. RESULTS: [Ga(III)/Lu(III)]-DOTA-PESIN showed good affinity to GRP and neuromedin B receptors but no affinity to BB3. [(67)Ga/(177)Lu]-DOTA-PESIN internalised rapidly into PC-3 cells whereas the efflux from PC-3 cells was relatively slow. In vivo experiments showed a high and specific tumour uptake and good retention of [(67)Ga/(177)Lu]-DOTA-PESIN. [(67)Ga/(177)Lu]-DOTA-PESIN highly accumulated in GRP receptor-expressing mouse pancreas. The uptake specificity was demonstrated by blocking tumour uptake and pancreas uptake. Fast clearance was found from blood and all non-target organs except the kidneys. High tumour-to-normal tissue ratios were achieved, which increased with time. PET imaging with [(68)Ga]-DOTA-PESIN was successful in visualising the tumour at 1 h post injection. Planar scintigraphic imaging showed that the (177)Lu-labelled peptide remained in the tumour even 3 days post injection. CONCLUSION: The newly designed ligands have high potential with regard to PET and SPECT imaging with (68/67)Ga and targeted radionuclide therapy with (177)Lu.